- Thermolysin is a thermostable zinc-dependent metalloprotease produced by Bacillus thermoproteolyticus. It catalyzes the hydrolysis of peptide bonds in proteins and peptides, with a strong preference for cleaving at the N-terminal side of hydrophobic amino acids such as leucine, isoleucine, valine, and phenylalanine. As a metalloendopeptidase, its catalytic activity relies on a zinc ion at the active site, which plays a critical role in stabilizing the transition state and polarizing the carbonyl group of the substrate’s scissile bond.
- One of the most distinguishing features of thermolysin is its thermal stability, with optimal activity at temperatures between 70–80 °C, and its ability to remain active at high temperatures where most other proteases denature. This makes it particularly useful in industrial applications, including protein hydrolysis in food processing, peptide synthesis, and biotechnology. It also tolerates a broad pH range and exhibits remarkable stability in organic solvents, further enhancing its utility in non-aqueous biocatalysis.
- Thermolysin’s structure has been well-characterized, revealing a compact fold stabilized by a network of disulfide bridges, hydrophobic interactions, and coordination with Zn²⁺. It belongs to the M4 family of metallopeptidases, often referred to as the thermolysin family. The enzyme employs a catalytic mechanism where the zinc ion and a glutamate residue activate a water molecule for nucleophilic attack on the peptide bond, assisted by other residues in a coordinated manner.
- In biochemical research, thermolysin is commonly used for limited proteolysis to analyze protein structure and domain organization. It is also applied in proteomics workflows, where its unique cleavage specificity complements that of trypsin, expanding peptide coverage during mass spectrometry analysis.
- In summary, thermolysin is a robust, heat-stable metalloprotease with broad applications in science and industry. Its unique biochemical properties and catalytic mechanism have made it a valuable tool in protein chemistry, enzymology, and industrial biotechnology.
